title: Naturally-occurring, dually-functional fusions between restriction endonucleases and regulatory proteins.
abstract: BACKGROUND
  Restriction-modification (RM) systems appear to play key roles in modulating gene flow among bacteria and archaea. Because the restriction endonuclease (REase) is potentially lethal to unmethylated new host cells, regulation to ensure pre-expression of the protective DNA methyltransferase (MTase) is essential to the spread of RM genes. This is particularly true for Type IIP RM systems, in which the REase and MTase are separate, independently-active proteins. A substantial subset of Type IIP RM systems are controlled by an activator-repressor called C protein. In these systems, C controls the promoter for its own gene, and for the downstream REase gene that lacks its own promoter. Thus MTase is expressed immediately after the RM genes enter a new cell, while expression of REase is delayed until sufficient C protein accumulates. To study the variation in and evolution of this regulatory mechanism, we searched for RM systems closely related to the well-studied C protein-dependent PvuII RM system. Unexpectedly, among those found were several in which the C protein and REase genes were fused.
  
  
  RESULTS
  The gene for CR.NsoJS138I fusion protein (nsoJS138ICR, from the bacterium Niabella soli) was cloned, and the fusion protein produced and partially purified. Western blots provided no evidence that, under the conditions tested, anything other than full-length fusion protein is produced. This protein had REase activity in vitro and, as expected from the sequence similarity, its specificity was indistinguishable from that for PvuII REase, though the optimal reaction conditions were different. Furthermore, the fusion was active as a C protein, as revealed by in vivo activation of a lacZ reporter fusion to the promoter region for the nsoJS138ICR gene.
  
  
  CONCLUSIONS
  Fusions between C proteins and REases have not previously been characterized, though other fusions have (such as between REases and MTases). These results reinforce the evidence for impressive modularity among RM system proteins, and raise important questions about the implications of the C-REase fusions on expression kinetics of these RM systems.
authors.affiliation: Department of Medical Microbiology & Immunology, College of Medicine and Life Sciences, University of Toledo, 3100 Transverse Drive, Toledo, OH 43614, USA. Robert.Blumenthal@utoledo.edu.
authors.forename: Jixiao
authors.initials: J
authors.lastname: Liang
authors.forename: Robert M
authors.initials: RM
authors.lastname: Blumenthal
mesh_terms: D041963:Bacteroidetes / Q000201:enzymology / Q000235:genetics / Q000378:metabolism*; D001483:Base Sequence; D015254:DNA Modification Methylases / Q000235:genetics / Q000378:metabolism; D004262:DNA Restriction Enzymes / Q000235:genetics / Q000378:metabolism; D015280:DNA Restriction-Modification Enzymes; D015257:DNA-Cytosine Methylases / Q000235:genetics / Q000378:metabolism; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic; D016415:Sequence Alignment; D014157:Transcription Factors / Q000235:genetics
chemical_list: D015280:DNA Restriction-Modification Enzymes; D014157:Transcription Factors; D015254:DNA Modification Methylases; C047084:DNA modification methylase PvuII; D015257:DNA-Cytosine Methylases; D004262:DNA Restriction Enzymes